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Fig. 3. Generation of Crb1^-/- mice and confocal images of mouse retinas. (A) Crb1 disrupted by insertion of the targeting vector. E1, exon 1; E2, Exon 2; pA, polyadenylation signal; PGK, phosphoglycerate kinase promoter; B, BamHI; RV, EcoRV; Bg, BglII. (B,C) Deletion of the exon encoding the N-terminal signal peptide prevents the production of Crb1 protein with C-terminal transmembrane and intracellular domains. (B) EcoRV Southern blot analysis using a 750 bp BglII-AccI fragment probe in the 3' flanking region. (C) Immunoprecipitation of Crb1, with AK7, from lysate of wild-type but not of the Crb1^-/- retina. As positive control 293/CRB1 cell lysates were used. Crb1 was stained using AK2. Asterisks indicate cross-reacting bands with AK2 in 293 cells. (D-G) Localization of Crb1 (red) in the OLM and staining of cone segments and pedicles by peanut agglutinin (PNA; green) in the retinas of wild-type (D) and Crb1^-/- (F) mice. Detail of the localization of Crb1 at the SAR for the wild-type (E) and Crb1^-/- (G) retina. Scale bars: 30 µm.

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